Novel objective classification of reactive microglia following hypoglossal axotomy using hierarchical cluster analysis

Abstract

A total of 136 microglia were intracellularly labeled and their morphological features were evaluated by 3D morphometric measurement. According to hierarchical cluster analysis, microglia were objectively categorized into four groups termed types I-IV. The validity of this classification was confirmed by principal component analysis and linear discriminant analysis. Type I microglia were found in sham-operated mice and in mice sacrificed 28 days (D28) after axotomy. The appearance of type I cells was similar to so-called ramified microglia in a resting state. Type II microglia were mainly seen in D14 mice, which exhibited small cell bodies with thin and short processes. Interestingly, none of the already-known morphological types of microglia seemed to be comparable to type II cells. We thus named type II microglia "small ramified" cells. Types III and IV microglia were mainly seen in D3 and D7 mice and their appearances were similar to hypertrophied and bushy cells, respectively. Proliferating cell nuclear antigen (PCNA), a mitosis marker, was almost exclusively expressed in D3 mice. On the other hand, voltage-dependent potassium channels (Kv1.3/1.5), neurotoxicity-related molecules, were most highly expressed in D14 mice. Increased expression of Kv1.3/1.5 in D14 mice was suppressed by minocycline treatment. These findings indicate that type II and III microglia may be involved in neurotoxicity and mitosis, respectively. Type IV microglial cells are assumed to be in the process of losing mitotic activity and gaining neurotoxicity. Our data also suggest that type II microglia can be a potential therapeutic target against neurodegenerative diseases.

Microglial proliferation in the brain of chronic alcoholics with hepatic encephalopathy

“…Using Iba1 stained thick sections allows visualization of the whole microglial cell in intricate detail to discriminate between ramified, activated and dystrophic microglia. The visible withdrawal and thickening of the processes in pHEs shown here is consistent with the morphological changes associated with microglial activation described by Yamada and Jinno in rodents (Yamada and Jinno 2013). Microglia are exceptionally dynamic, plastic cells, with a wide range of physiological functions beyond their phagocytic role such as monitoring the state of synapses (Wake et al 2009).…”

Comparative morphometric analysis of microglia in the spinal cord of SOD1G93A transgenic mouse model of amyotrophic lateral sclerosis

“…Chronic administration of IDPN is thus considered to partially mimic an early event in the pathogenesis of ALS via inhibition of axonal flow of motoneurons (Williamson & Cleveland, ; Wang et al ., ). The general appearance of type R1 microglia is similar to ‘type II’ microglia, which we have found in the hypoglossal nucleus 14 days after axotomy (Yamada & Jinno, ). Note that axonal degeneration begins 3–4 days after the injury, and is completed after 12–14 days (Rotshenker, ).…”

Differential activation of neuronal and glial STAT3 in the spinal cord of the SOD1G93A mouse model of amyotrophic lateral sclerosis

“…No bands of equivalent size are observed in analyses of brain samples of Kv1.3‐knockout mice (manufacturer's technical information). This antibody's staining pattern of activated microglia was similar to those of previous studies (Yamada & Jinno, ; Rangaraju et al ., ).…”

“…No bands of equivalent size are observed in Kv1.5‐untransfected COS cells (manufacturer's technical information). This antibody's staining pattern of activated microglia was identical to those reported previously (Jou et al ., ; Yamada & Jinno, ).…”